The Molecular Diagnostics Section is currently the only CLIA and College of American Pathology approved clinical laboratory within the NCI certified for performing molecular oncology testing on pathology materials from NIH patients. In 2017 the molecular diagnostics laboratory will process a projected 2150 unique clinical samples from NCI/NIH patients, and performed 6885 tests. The laboratory utilizes a variety of technologies to perform these assays including conventional DNA PCR, RT-PCR, qPCR, droplet digital PCR, allele suppression PCR, capillary electrophoresis, pyrosequencing, bisulfite pyrosequencing, and small panel next generation sequencing (NGS). The assays we perform include tests to identify B and T-cell clonality, lymphoma specific translocations (e.g., BCL1/IG, BCL2/IG), translocations associated with pediatric sarcomas (e.g., EWS-FLI-1, EWS-ERG, EWS-WT-1, PAX-7/FKHD, PAX-3/FKHD, SYT-SSX-1, and SYT-SSX-2), cancer associated viruses (e.g., EBV, HTLV, HHV-8), and mutations associated with a variety of cancers (e.g., EGFR, KRAS, NRAS, BRAF, PIK3CA, IDH1/2, AKT, MYD88, ERBB2, STAT3, and STAT5b). Quantitative PCR is performed for HTLV 1/2 to follow viral load in ATL clinical trials, and for the EGFRvIII to assess expression of this cancer-specific transcript for clinical trial eligibility. MGMT methylation analysis is performed using bisulfite pyrosequencing to provide predictive information regarding response to temozolomide in the glioblastoma setting. The laboratory has developed a custom CNS NGS panel to assist in the classification of CNS cancers referred to the NeuroOncology Branch (NOB). It is now an integral diagnostic component of the NOB's natural history protocol. The laboratory validated and replaced its standard 50 gene NGS panel with the NCI MATCH program equivalent Oncomine Comprehensive Assay v3 for use in clinical tumor profiling. The laboratory's developmental research effort has been focused on the application of next generation sequencing (NGS) technologies to assist in cancer genotyping and diagnostics. As previously mentioned, the laboratory developed a primary brain tumor panel to assist with diagnosis and classification of central nervous system cancers as part of a major collaboration with the Neuro-Oncology Branch of CCR. This panel has already been shown to have clinical utility. Over 10% of the first 300 cases studied have had diagnoses changed as a result of the information provided by this panel. The laboratory has also initiated a program in the use of circulating tumor DNA (ctDNA) as a biomarker for disease response and early detection. These two efforts (NGS and ctDNA) are linked as the genotyping is not only used to identify potential therapeutic targets, but it is also used to identify targets for the circulating DNA (ctDNA) studies. Early studies targeting BRAF V600E ctDNA in melanoma patients (collaboration with Dr. S. Rosenberg) indicated the potential use of this technology in predicting responses to cellular immunotherapy, and recurrence. We have expanded our initial studies with Dr. Rosenberg in melanoma to a wide range of other TIL treated tumors (breast, ovary, colon) targeting patient specific exome and transcriptome sequence identified targets, with similar results. We have initiated new studies with other investigators in diverse cancer subtypes including uveal melanoma, lung, bladder, and pancreatic cancer. Early results with Dr. Guha in lung cancer has shown the sensitivity of the technology in tracing disease activity, and identifying treatment failures weeks to months before progressive disease became clinically evident. We have continued to study rare hematopoietic cancers in collaboration with Dr. Elaine Jaffe and are currently investigating the molecular biology of Histiocytic Sarcoma taking advantage of the samples collected and archived by Dr. Jaffe in her consult practice, and have identified novel subgroups of these rare cancers based on gene expression, disease site and mutational status. The molecular diagnostics laboratory also supports translational research of NCI and NIH researchers. Among the NCI and NIH investigators and clinicians that have utilized the laboratory's resources in 2017-18 are: Dr. A. Apolo (NCI), Dr. Jeffrey Cohen (NIAID), Dr. William Gahl (NHGRI), Dr. Mark Gilbert (NCI), Dr. Udayan Guha, Dr. Raffit Hassan (NCI), Dr. Steven Holland (NIAID), Dr. Elaine Jaffe (NCI), Dr. Amy Klion (NHLBI) Dr. Robert Kreitman (NCI) Dr. Markku Mietinnen (NCI), Dr. Stefania Pittaluga (NCI), Dr. Martha Quezado (NCI), Dr. Arun Rajan (NCI), Dr. Koneti Rao (NIAID), Dr. Steven Rosenberg (NCI), Dr. Udo Rudoff (NCI), Dr. Helen Su (NHLBI), Dr. Anish Thomas, Dr. Gulbu Uzel (NIAID), Dr. Katherine Warren (NCI), Dr. Wyndham Wilson (NCI), Dr. James Yang (NCI), Dr. Neil Young (NHLBI).